Luteinizing hormone-releasing hormone (LHRH) is a neurotransmitter produced by the hypothalamus which stabilizes the secretion of luteinizing hormone (LHRH) and follicle-stimulating hormone (FSH) from the pituitary, which in turn stimulates the synthesis of steroid hormones, such as testosterone, from the gonads. Many LHRH peptide analogs (e.g., agonists and antagonists) are currently sold for the treatment of endometriosis, prostate cancer, precocious puberty, and other hormonally mediated disorders. The synthesis of such LHRH analogs are difficult and expensive.
Solid phase peptide synthesis was introduced in 1963 with the intent to overcome many of the intermediate purification problems associated with solution phase peptide synthesis. Stewart, et al., Solid Phase Peptide Synthesis (Pierce Chemical Co., 2d ed.) 1984. During solid phase synthesis, amino acids are assembled (i.e., coupled) into a peptide of any desired sequence while one end of the chain (i.e., the C-terminus) is anchored to an insoluble support. Once the desired sequence has been linked together on the support, the peptide is then detached (i.e., cleaved) from the support.
U.S. Pat. No. 4,010,125 (1977) describes the synthesis of [D-Trp6]-LHRH in which the side chains of four of the ten residues of the LHRH agonist are protected (e.g., L-histidyl(tosyl), seryl(benzyl), tyrosyl(2,6-di-Cl-Bzl), and L-arginyl(tosyl)). However, Coy, et al., Int. J. Peptide Protein Res. 14:359 (1979) later reported the synthesis of a number of other LHRH analogs in which minimal side chain protection, e.g., only salt protection of arginine (i.e., Arg.HCl), produced superior yield to the corresponding protected synthesis. The present invention is directed to the discovery that, despite the trend in the art to not protect the side chains of amino acids in the synthesis of LHRH analogs, the protection of the tryptophan residue during solid phase synthesis does indeed improve the yield of LHRH analogs having a tryptophan residue (e.g., [D)-Trp6]LHRH).